1. Field of the Invention
The present invention relates to a bio-silica chip comprising a silica-binding protein and a fabrication method thereof, and more particularly to a bio-silica chip in which a fusion protein of a silica-binding protein and a probe protein is immobilized on a chip comprising a silica layer, a fabrication method thereof, and a method for detecting interactions between biomaterials using the bio-silica chip.
2. Background of the Related Art
Recently, the functions of 10,000 genes among 100,000 predicted human genes were identified by the human genome project, and most of such genes are known to have direct connections to diseases. Also, because most diseases occur at the protein level rather than at the gene level, more than 95% of drugs, which have been developed to date or are being developed, target proteins. Thus, a protein sensor chip is a core technology in researches to find out the functions of biomolecules interacting specifically with a certain protein and to develop a method for treating and preventing diseases which was impossible to treat using classic methods, on the basis of the data obtained from protein function analysis and network analysis. Also, the protein sensor chip can selectively analyze small amounts of materials and can perform real-time measurement of the materials, and thus is widely used not only for the research of basic science, but also for the measurement of pollutants and the diagnosis of drugs or viruses.
A typical chip among biosensors including protein sensor chips is silica chips. In technologies on the analysis of protein-protein, DNA-RNA or carbohydrate-protein interactions using silica chips, which have been performed to date, a probe protein is immobilized on an affinity tag (Ni-NTA, Streptavidin, GST, etc.) or a silica chip surface after chemically modifying the silica chip surface with, for example, amine, ligand thiol or aldehyde, or by forming a self-assembly monolayer of a ligand having functional groups attached thereto. Alternatively, a probe protein is site-specifically immobilized using a technology of forming a uniform and stable monolayer of a biomolecule to control the orientation of the biomolecule at the molecular level. Using the immobilized probe protein, protein-protein interactions are analyzed using surface plasmon resonance (SPR) or the like (Hentz et al., Anal. Chem., 68:3939, 1996; Kukar et al, Anal. Biochem., 306:50, 2002; Hall D., Anal Biochem., 288:109, 2001; Canziani, G. et al, Methods, 19:253, 1999; Brockman, J. M. et al, JACS, 121:8044, 1999; Nelson, B. P. et al, Anal Chem., 73:1, 2001; Jordan, C. E. et al, Anal Chem., 69:4939, 1997; Goodrich, T. T. et al, JACS, 126:4086, 2004).
However, although various biosensor technologies using silica chips as described above were developed, the technology for treating the surface of silica chips has great disadvantages in which a method for chemically treating the chip surface is complicated, and non-specific protein binding occurs. Also, it is difficult for this technology to be practically used, because proteins have weak binding ability and can be influenced by many chemical substances, thus causing many limitations in detecting protein-protein interactions. Furthermore, because high purity is required to immobilize the protein on the silica chip, a complicated purification process must be included, thus reducing economic efficiency Accordingly, there has been a need to develop a novel chip overcoming such shortcomings.
Moreover, the development of technology capable of selectively and stably immobilizing biomaterials on biosensor chips is required for the development of biosensors. Conventional methods for immobilizing biomaterials on biosensor chips are broadly classified into the following four methods: a method of physically or chemically adsorbing biomaterials onto the surface of the sensor chip, a method of covalently bonding biomaterials to the senor chip surface, a method of capturing biomaterials onto membranes, matrices, polymers, etc., and a method of immobilizing biomaterials on the sensor chip surface by inducing crosslinking between the biomaterials. Thus, there has been a need to develop a novel immobilization method different from such methods, which can selectively and stably immobilize biomaterials to the sensor chip surface.
Meanwhile, since it was known that, among silica-binding proteins (SBPs), a silica-binding protein consisting of 12 amino acids binds selectively to the surface of silica (Brown, S., Nature Biotechnol., 15:269, 1997; Naik, R. R. et al., J. Nanosci. Nanotechnol., 2:1, 2002), studies on SBP have been conducted. However, the exact mechanism by which on how SBP recognizes and binds to the silica surface is not yet found. It is predicted that the hydrophobic portion of the silica surface has a specific interaction with the hydrophobic region of the SBP sequence on the basis of molecular recognition (Braun, R., et al, J. Biomater. Sci., 13:747, 2002).
Accordingly, the present inventors have made many efforts to provide a novel bio-silica chip overcoming the problems of the conventional silica chips, which can specifically and stably immobilize a probe protein, and is economical. As a result, the present inventors have isolated a novel silica-binding protein and found that a fusion protein of the isolated silica-binding protein and a probe protein is position-specifically immobilized on a silica chip, and the immobilized fusion protein can bind specifically to a target, thus making it possible to efficiently analyze the interaction, thereby completing the present invention.